@gmod/bam 8.5.1 → 8.7.0

package/dist/mismatches.js

@@ -0,0 +1,404 @@
+"use strict";
+Object.defineProperty(exports, "__esModule", { value: true });
+exports.MISMATCH_HARD_CLIP = exports.MISMATCH_SOFT_CLIP = exports.MISMATCH_REF_SKIP = exports.MISMATCH_DELETION = exports.MISMATCH_INSERTION = exports.MISMATCH_SUBST = void 0;
+exports.forEachMismatchNumeric = forEachMismatchNumeric;
+const cigar_ts_1 = require("./cigar.js");
+const reference_ts_1 = require("./reference.js");
+const seqAlphabet_ts_1 = require("./seqAlphabet.js");
+/**
+ * The kind of difference a {@link Mismatch} reports, as the CIGAR-style char
+ * code it corresponds to. `@gmod/cram`'s `forEachMismatch` emits the same
+ * values (its `RF_*` constants), so one `switch` handles records from either
+ * format.
+ *
+ * Note these are char codes and NOT the packed-CIGAR op numbers in `cigar.ts`
+ * — `MISMATCH_DELETION` is 68 (`'D'`), where `CIGAR_DEL` is 2.
+ */
+exports.MISMATCH_SUBST = 88; // 'X'
+exports.MISMATCH_INSERTION = 73; // 'I'
+exports.MISMATCH_DELETION = 68; // 'D'
+exports.MISMATCH_REF_SKIP = 78; // 'N'
+exports.MISMATCH_SOFT_CLIP = 83; // 'S'
+exports.MISMATCH_HARD_CLIP = 72; // 'H'
+// M and = both mean "aligned to the reference", differing only in whether the
+// file promises the bases match. Which is exactly the distinction MD (or the
+// reference) exists to settle, so the walk treats them identically.
+const CIGAR_M_EQ_MASK = (1 << cigar_ts_1.CIGAR_MATCH) | (1 << cigar_ts_1.CIGAR_EQUAL);
+// A clip has no bases to report — its length travels in `clipLength`, which is
+// what every consumer reads.
+const NO_BASES = '';
+function reportRefMismatch(callback, refPos, seqNibble, qual, refNibble) {
+    callback(exports.MISMATCH_SUBST, refPos, 1, seqAlphabet_ts_1.SEQRET_DECODER[seqNibble], qual, reference_ts_1.CHAR_CODE_FROM_NIBBLE[refNibble], 0);
+}
+// One base compared on its own: the ends of a run whose length or alignment
+// leaves it without a partner in the same byte.
+function compareRefBase(callback, numericSeq, qual, ref, seqIdx, refIdx, refPos) {
+    const seqNibble = (numericSeq[seqIdx >> 1] >> ((1 - (seqIdx & 1)) << 2)) & 0xf;
+    const refNibble = (0, reference_ts_1.referenceNibble)(ref, refIdx);
+    if (seqNibble !== refNibble) {
+        reportRefMismatch(callback, refPos, seqNibble, qual ? qual[seqIdx] : -1, refNibble);
+    }
+}
+/**
+ * The mismatch walk over a record's already-packed fields, for callers holding
+ * BAM's arrays without a {@link BamRecord} around them — a SAM parser, or a
+ * worker that was handed the typed arrays rather than the record.
+ * `record.forEachMismatch()` is this function with the record's own fields
+ * filled in, and is what most code should call.
+ *
+ * Nothing here allocates per difference except the `bases` string, and that
+ * only for substitutions and insertions.
+ *
+ * @param cigar packed CIGAR ops, i.e. `record.NUMERIC_CIGAR`
+ * @param numericSeq 4-bit packed SEQ, i.e. `record.NUMERIC_SEQ`
+ * @param seqLength bases in `numericSeq`; 0 for a read stored with `SEQ` of `*`
+ * @param md the MD tag's bytes (`record.NUMERIC_MD`), or undefined. Preferred
+ *   over `ref` when present, being both cheaper and what the aligner asserted
+ * @param qual per-base quality scores, or null/undefined when the read has none
+ * @param ref reference bases to compare against when there is no MD tag. Bases
+ *   the region does not cover are left uncompared
+ * @param refStart the record's own 0-based reference start
+ * @param windowStart 0-based half-open reference window to report within, in
+ *   ABSOLUTE reference coordinates whatever `origin` is. Pass
+ *   `-Infinity`/`Infinity` for the whole read; a viewport that clips the walk
+ *   is what keeps a chromosome-spanning contig alignment from being walked in
+ *   full for every screenful
+ * @param windowEnd
+ * @param origin what reported positions are relative to: 0 for reference
+ *   positions, `refStart` for read-relative ones. See
+ *   {@link MismatchOptions.origin}
+ * @param callback `(code, refPos, length, bases, qual, refBaseCode, clipLength)`
+ */
+function forEachMismatchNumeric(cigar, numericSeq, seqLength, md, qual, ref, refStart, windowStart, windowEnd, origin, callback) {
+    // The walk runs in read-relative offsets — `roffset` reference bases and
+    // `soffset` read bases consumed — and converts back at the callback, so the
+    // window is converted once here rather than per op.
+    //
+    // Clamped to int32 rather than left at the ±Infinity an unwindowed walk
+    // passes, because every op of the walk compares an offset against these and
+    // Infinity makes each of those a Float64 comparison. BAM positions are int32,
+    // so this cannot narrow a window anyone can express.
+    const winLo = windowStart <= -0x80000000 ? -0x80000000 : windowStart - refStart;
+    const winHi = windowEnd >= 0x7fffffff ? 0x7fffffff : windowEnd - refStart;
+    // What an offset is emitted as. Folding `origin` in here rather than at the
+    // callbacks keeps the cost of having the option at zero: every emit already
+    // added `refStart`, and now adds this instead.
+    const outBase = refStart - origin;
+    // Fast path for reads with no sequence (e.g. secondary alignments with SEQ='*')
+    if (seqLength === 0) {
+        let roffset = 0;
+        for (let i = 0, l = cigar.length; i < l; i++) {
+            // `roffset` only grows, and nothing at or past the window's right edge is
+            // reported, so the rest of the CIGAR cannot produce anything (see the
+            // same check in the main loop)
+            if (roffset >= winHi) {
+                break;
+            }
+            const packed = cigar[i];
+            const len = packed >>> 4;
+            const op = packed & 0xf;
+            // X consumes the reference exactly like M/=; without a sequence there are
+            // no bases to report for it, but it still has to advance — leaving it out
+            // shifted every later op left by the total X length, which is every indel
+            // in an --eqx CIGAR walked without a sequence.
+            if ((1 << op) & CIGAR_M_EQ_MASK || op === cigar_ts_1.CIGAR_DIFF) {
+                roffset += len;
+            }
+            else if (op === cigar_ts_1.CIGAR_INS) {
+                if (roffset >= winLo && roffset < winHi) {
+                    callback(exports.MISMATCH_INSERTION, outBase + roffset, 0, '', -1, 0, len);
+                }
+            }
+            else if (op === cigar_ts_1.CIGAR_DEL) {
+                if (roffset < winHi && roffset + len > winLo) {
+                    callback(exports.MISMATCH_DELETION, outBase + roffset, len, NO_BASES, -1, 0, 0);
+                }
+                roffset += len;
+            }
+            else if (op === cigar_ts_1.CIGAR_REF_SKIP) {
+                if (roffset < winHi && roffset + len > winLo) {
+                    callback(exports.MISMATCH_REF_SKIP, outBase + roffset, len, NO_BASES, -1, 0, 0);
+                }
+                roffset += len;
+            }
+            else if (op === cigar_ts_1.CIGAR_SOFT_CLIP) {
+                if (roffset >= winLo && roffset < winHi) {
+                    callback(exports.MISMATCH_SOFT_CLIP, outBase + roffset, 0, NO_BASES, -1, 0, len);
+                }
+            }
+            else if (op === cigar_ts_1.CIGAR_HARD_CLIP) {
+                if (roffset >= winLo && roffset < winHi) {
+                    callback(exports.MISMATCH_HARD_CLIP, outBase + roffset, 0, NO_BASES, -1, 0, len);
+                }
+            }
+        }
+        return;
+    }
+    const mdLength = md?.length ?? 0;
+    const hasQual = !!qual;
+    const hasMD = md && mdLength > 0;
+    // Where the reference region sits in read-relative offsets, and the window
+    // narrowed to it. Only base COMPARISON is bounded by the region — an indel or
+    // a clip needs no reference base, so a region covering part of a read still
+    // reports every one of them.
+    //
+    // Ternaries rather than Math.max/Math.min, which matters more than it looks:
+    // an unwindowed walk has `winLo`/`winHi` at ±Infinity, so Math.min would hand
+    // back a Float64 and `cmpHi` would carry that all the way into `jHi`, the
+    // bound of the innermost (two-bases-per-byte) loop. This way an unwindowed
+    // walk keeps that bound a Smi, which measured ~8% of the reference path on
+    // dense short reads.
+    let refOffset = 0;
+    let cmpLo = 0;
+    let cmpHi = 0;
+    if (ref !== undefined) {
+        refOffset = refStart - ref.start;
+        const refLo = -refOffset;
+        const refHi = ref.length - refOffset;
+        cmpLo = winLo > refLo ? winLo : refLo;
+        cmpHi = winHi < refHi ? winHi : refHi;
+    }
+    let roffset = 0;
+    let soffset = 0;
+    let mdIdx = 0;
+    let mdMatchRemaining = 0;
+    if (hasMD) {
+        while (mdIdx < mdLength) {
+            const c = md[mdIdx];
+            if (c >= 48 && c <= 57) {
+                mdMatchRemaining = mdMatchRemaining * 10 + (c - 48);
+                mdIdx++;
+            }
+            else {
+                break;
+            }
+        }
+    }
+    for (let i = 0, l = cigar.length; i < l; i++) {
+        // Stop at the window's right edge rather than walking the rest of the
+        // CIGAR: `roffset` only grows, and every report below needs
+        // `roffset < winHi`, so nothing past here can produce anything. Matters for
+        // a whole chromosome stored as one BAM read, whose CIGAR runs to millions
+        // of ops for a screenful of them — and costs one compare per op otherwise,
+        // since an unwindowed walk has `winHi` at +Infinity.
+        if (roffset >= winHi) {
+            break;
+        }
+        const packed = cigar[i];
+        const len = packed >>> 4;
+        const op = packed & 0xf;
+        if ((1 << op) & CIGAR_M_EQ_MASK) {
+            if (hasMD) {
+                let remaining = len;
+                let localOffset = 0;
+                while (remaining > 0) {
+                    if (mdMatchRemaining >= remaining) {
+                        mdMatchRemaining -= remaining;
+                        localOffset += remaining;
+                        remaining = 0;
+                    }
+                    else {
+                        localOffset += mdMatchRemaining;
+                        remaining -= mdMatchRemaining;
+                        mdMatchRemaining = 0;
+                        if (mdIdx < mdLength && md[mdIdx] >= 65 && md[mdIdx] <= 90) {
+                            const pos = roffset + localOffset;
+                            if (pos >= winLo && pos < winHi) {
+                                const seqIdx = soffset + localOffset;
+                                const sb = numericSeq[seqIdx >> 1];
+                                const nibble = (sb >> ((1 - (seqIdx & 1)) << 2)) & 0xf;
+                                callback(exports.MISMATCH_SUBST, outBase + pos, 1, seqAlphabet_ts_1.SEQRET_DECODER[nibble], hasQual ? qual[seqIdx] : -1, md[mdIdx], 0);
+                            }
+                            mdIdx++;
+                            localOffset++;
+                            remaining--;
+                            mdMatchRemaining = 0;
+                            while (mdIdx < mdLength) {
+                                const c = md[mdIdx];
+                                if (c >= 48 && c <= 57) {
+                                    mdMatchRemaining = mdMatchRemaining * 10 + (c - 48);
+                                    mdIdx++;
+                                }
+                                else {
+                                    break;
+                                }
+                            }
+                        }
+                        else {
+                            break;
+                        }
+                    }
+                }
+            }
+            else if (ref) {
+                // Only compare bases whose reference position falls in the window AND
+                // in the region. A whole-chromosome contig has M ops totalling ~250M
+                // bases; without this clip every one is compared (and `ref` would have
+                // to cover the whole chromosome).
+                const jLo = cmpLo > roffset ? cmpLo - roffset : 0;
+                const jHi = cmpHi < roffset + len ? cmpHi - roffset : len;
+                let j = jLo;
+                // A leading odd read index has no partner in its byte, so compare it
+                // alone and let the pair loop start on a byte boundary.
+                if (j < jHi && (soffset + j) & 1) {
+                    compareRefBase(callback, numericSeq, qual, ref, soffset + j, refOffset + roffset + j, outBase + roffset + j);
+                    j++;
+                }
+                // Two bases per byte load and compare. `even`/`odd` differ only in
+                // which reference parity they put on a byte boundary, so one of them
+                // always lines up with the read's packed sequence; the nibbles are
+                // unpacked only for the byte that actually differs.
+                const refIdx = refOffset + roffset + j;
+                const refPairs = refIdx & 1 ? ref.odd : ref.even;
+                let refByte = (refIdx + (refIdx & 1)) >> 1;
+                let seqByte = (soffset + j) >> 1;
+                for (; j + 1 < jHi; j += 2, refByte++, seqByte++) {
+                    const seqPair = numericSeq[seqByte];
+                    const refPair = refPairs[refByte];
+                    if (seqPair !== refPair) {
+                        const seqHi = (seqPair >> 4) & 0xf;
+                        const refHi = (refPair >> 4) & 0xf;
+                        if (seqHi !== refHi) {
+                            reportRefMismatch(callback, outBase + roffset + j, seqHi, hasQual ? qual[soffset + j] : -1, refHi);
+                        }
+                        const seqLo = seqPair & 0xf;
+                        const refLo = refPair & 0xf;
+                        if (seqLo !== refLo) {
+                            reportRefMismatch(callback, outBase + roffset + j + 1, seqLo, hasQual ? qual[soffset + j + 1] : -1, refLo);
+                        }
+                    }
+                }
+                if (j < jHi) {
+                    compareRefBase(callback, numericSeq, qual, ref, soffset + j, refOffset + roffset + j, outBase + roffset + j);
+                }
+            }
+            soffset += len;
+            roffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_INS) {
+            if (roffset >= winLo && roffset < winHi) {
+                // Optimized insertion base extraction - avoid string concat for common
+                // cases
+                let insertedBases;
+                if (len === 1) {
+                    // Single base insertion - most common case
+                    const sb = numericSeq[soffset >> 1];
+                    const nibble = (sb >> ((1 - (soffset & 1)) << 2)) & 0xf;
+                    insertedBases = seqAlphabet_ts_1.SEQRET_DECODER[nibble];
+                }
+                else if (len === 2) {
+                    // Two base insertion - second most common
+                    const seqIdx0 = soffset;
+                    const sb0 = numericSeq[seqIdx0 >> 1];
+                    const nibble0 = (sb0 >> ((1 - (seqIdx0 & 1)) << 2)) & 0xf;
+                    const seqIdx1 = soffset + 1;
+                    const sb1 = numericSeq[seqIdx1 >> 1];
+                    const nibble1 = (sb1 >> ((1 - (seqIdx1 & 1)) << 2)) & 0xf;
+                    insertedBases = seqAlphabet_ts_1.SEQRET_DECODER[nibble0] + seqAlphabet_ts_1.SEQRET_DECODER[nibble1];
+                }
+                else {
+                    const bases = new Array(len);
+                    for (let j = 0; j < len; j++) {
+                        const seqIdx = soffset + j;
+                        const sb = numericSeq[seqIdx >> 1];
+                        const nibble = (sb >> ((1 - (seqIdx & 1)) << 2)) & 0xf;
+                        bases[j] = seqAlphabet_ts_1.SEQRET_DECODER[nibble];
+                    }
+                    insertedBases = bases.join('');
+                }
+                callback(exports.MISMATCH_INSERTION, outBase + roffset, 0, insertedBases, -1, 0, len);
+            }
+            soffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_DEL) {
+            if (roffset < winHi && roffset + len > winLo) {
+                callback(exports.MISMATCH_DELETION, outBase + roffset, len, NO_BASES, -1, 0, 0);
+            }
+            // MD spells a deletion as ^ then the deleted reference bases, which are
+            // not a difference this reports (the D above already is) but do have to
+            // be stepped over, or every later position in the read reads off the
+            // wrong part of the tag.
+            // eslint-disable-next-line @typescript-eslint/no-confusing-non-null-assertion
+            if (hasMD && mdIdx < mdLength && md[mdIdx] === 94) {
+                mdIdx++;
+                while (mdIdx < mdLength && md[mdIdx] >= 65) {
+                    mdIdx++;
+                }
+                mdMatchRemaining = 0;
+                while (mdIdx < mdLength) {
+                    const c = md[mdIdx];
+                    if (c >= 48 && c <= 57) {
+                        mdMatchRemaining = mdMatchRemaining * 10 + (c - 48);
+                        mdIdx++;
+                    }
+                    else {
+                        break;
+                    }
+                }
+            }
+            roffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_REF_SKIP) {
+            if (roffset < winHi && roffset + len > winLo) {
+                callback(exports.MISMATCH_REF_SKIP, outBase + roffset, len, NO_BASES, -1, 0, 0);
+            }
+            roffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_DIFF) {
+            for (let j = 0; j < len; j++) {
+                const seqIdx = soffset + j;
+                const sb = numericSeq[seqIdx >> 1];
+                const nibble = (sb >> ((1 - (seqIdx & 1)) << 2)) & 0xf;
+                // An X op says a substitution is here without saying what it replaces,
+                // so the reference base comes from MD or the region — and is 0, i.e.
+                // unknown, when the read carries neither.
+                let refBaseCode = 0;
+                if (hasMD) {
+                    if (mdMatchRemaining === 0 && mdIdx < mdLength && md[mdIdx] >= 65) {
+                        refBaseCode = md[mdIdx];
+                        mdIdx++;
+                        mdMatchRemaining = 0;
+                        while (mdIdx < mdLength) {
+                            const c = md[mdIdx];
+                            if (c >= 48 && c <= 57) {
+                                mdMatchRemaining = mdMatchRemaining * 10 + (c - 48);
+                                mdIdx++;
+                            }
+                            else {
+                                break;
+                            }
+                        }
+                    }
+                    else if (mdMatchRemaining > 0) {
+                        mdMatchRemaining--;
+                    }
+                }
+                else if (ref) {
+                    const refIdx = refOffset + roffset + j;
+                    if (refIdx >= 0 && refIdx < ref.length) {
+                        refBaseCode = reference_ts_1.CHAR_CODE_FROM_NIBBLE[(0, reference_ts_1.referenceNibble)(ref, refIdx)];
+                    }
+                }
+                const pos = roffset + j;
+                if (pos >= winLo && pos < winHi) {
+                    callback(exports.MISMATCH_SUBST, outBase + pos, 1, seqAlphabet_ts_1.SEQRET_DECODER[nibble], hasQual ? qual[seqIdx] : -1, refBaseCode, 0);
+                }
+            }
+            soffset += len;
+            roffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_SOFT_CLIP) {
+            if (roffset >= winLo && roffset < winHi) {
+                callback(exports.MISMATCH_SOFT_CLIP, outBase + roffset, 0, NO_BASES, -1, 0, len);
+            }
+            soffset += len;
+        }
+        else if (op === cigar_ts_1.CIGAR_HARD_CLIP) {
+            if (roffset >= winLo && roffset < winHi) {
+                callback(exports.MISMATCH_HARD_CLIP, outBase + roffset, 0, NO_BASES, -1, 0, len);
+            }
+        }
+        // P (padding) consumes neither read nor reference and is not a difference
+    }
+}
+//# sourceMappingURL=mismatches.js.map

package/dist/mismatches.js.map

@@ -0,0 +1 @@
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package/dist/record.d.ts

@@ -1,3 +1,5 @@
+import type { Mismatch, MismatchCallback, MismatchOptions } from './mismatches.ts';
+import type { PackedReference } from './reference.ts';
 export type NumericCigar = Uint32Array | Int32Array | number[];
 export default class BamRecord {
     fileOffset: number;
@@ -11,6 +13,7 @@ export default class BamRecord {
     private _cachedNumericCigar?;
     private _cachedNUMERIC_MD?;
     private _cachedSeqStart?;
+    private _reference?;
     constructor(byteArray: Uint8Array, start: number, end: number, fileOffset: number, dataView: DataView);
     get byteArray(): Uint8Array<ArrayBufferLike>;
     get flags(): number;
@@ -79,5 +82,48 @@ export default class BamRecord {
     get next_pos(): number;
     get template_length(): number;
     seqAt(idx: number): string | undefined;
+    /**
+     * Bind reference bases for this read to resolve its substitutions against,
+     * for a read with no MD tag. `BamFile` calls this for you when it was given a
+     * `fetchReferenceSequence`; call it yourself when you have the bases from
+     * somewhere else.
+     *
+     * **`ref` must cover the whole read**, and this throws if it does not. The
+     * reason is that records are cached and shared between queries (ADR 0006), so
+     * a binding that varied by query would make one query's reads answer out of
+     * another's region. A region covering the read is the same data whichever
+     * query fetched it, so binding it is safe; a partial one is not, and the
+     * per-call `forEachMismatch(cb, {ref})` is where that belongs.
+     */
+    setReference(ref: PackedReference | undefined): void;
+    /** the region {@link setReference} bound, if any */
+    get reference(): PackedReference | undefined;
+    /**
+     * Report each difference between this read and the reference —
+     * substitutions, insertions, deletions, reference skips and clips — without
+     * allocating an object per difference. This is the intended way to read a
+     * record's differences; deriving them from `CIGAR` and `MD` yourself takes
+     * rather more of the format to interpret correctly (see {@link Mismatch}).
+     *
+     * Substitutions come from the MD tag when the read has one, and otherwise
+     * from comparing SEQ against reference bases — which have to come from
+     * {@link setReference}, `opts.ref`, or the file's `fetchReferenceSequence`.
+     * With neither MD nor reference, indels and clips are still reported in full
+     * and substitutions are not reported at all: nothing in the record says where
+     * they are.
+     *
+     * @param callback called as
+     *   `(code, refPos, length, bases, qual, refBaseCode, clipLength)`
+     * @param opts optional reference window to restrict to, an `origin` the
+     *   reported positions are relative to, and an optional per-call reference
+     *   region; see {@link MismatchOptions}
+     */
+    forEachMismatch(callback: MismatchCallback, opts?: MismatchOptions): void;
+    /**
+     * The same differences {@link forEachMismatch} reports, as an array of
+     * {@link Mismatch} objects. Convenient; allocates one object per difference,
+     * so the callback form is the one to reach for on a hot path.
+     */
+    getMismatches(opts?: MismatchOptions): Mismatch[];
     toJSON(): Record<string, unknown>;
 }

package/dist/record.js

@@ -5,9 +5,9 @@ var __importDefault = (this && this.__importDefault) || function (mod) {
 Object.defineProperty(exports, "__esModule", { value: true });
 const cigar_ts_1 = require("./cigar.js");
 const constants_ts_1 = __importDefault(require("./constants.js"));
-const SEQRET = '=ACMGRSVTWYHKDBN';
-const SEQRET_DECODER = SEQRET.split('');
-const SEQRET_CODES = Uint8Array.from(SEQRET, c => c.charCodeAt(0));
+const mismatches_ts_1 = require("./mismatches.js");
+const reference_ts_1 = require("./reference.js");
+const seqAlphabet_ts_1 = require("./seqAlphabet.js");
 // Both bases of a SEQ byte, precomputed for all 256 bytes so decoding advances a
 // byte at a time. Two forms because `seq` has two strategies (see below): packed
 // ASCII codes for a single Uint16Array store, and the 2-char string to append.
@@ -17,13 +17,13 @@ const SEQRET_PAIR_CODES = new Uint16Array(256);
 const SEQRET_PAIR_STRINGS = new Array(256);
 for (let hi = 0; hi < 16; hi++) {
     for (let lo = 0; lo < 16; lo++) {
-        const h = SEQRET_CODES[hi];
-        const l = SEQRET_CODES[lo];
+        const h = seqAlphabet_ts_1.SEQRET_CODES[hi];
+        const l = seqAlphabet_ts_1.SEQRET_CODES[lo];
         SEQRET_PAIR_CODES[(hi << 4) | lo] = LITTLE_ENDIAN
             ? h | (l << 8)
             : (h << 8) | l;
         SEQRET_PAIR_STRINGS[(hi << 4) | lo] =
-            SEQRET_DECODER[hi] + SEQRET_DECODER[lo];
+            seqAlphabet_ts_1.SEQRET_DECODER[hi] + seqAlphabet_ts_1.SEQRET_DECODER[lo];
     }
 }
 // Read length at which building a byte buffer and calling TextDecoder once
@@ -325,6 +325,7 @@ class BamRecord {
     _cachedNumericCigar;
     _cachedNUMERIC_MD;
     _cachedSeqStart;
+    _reference;
     // Positional rather than an options object, because every argument is
     // unpacked into a field immediately and nothing keeps the wrapper. The
     // options form allocated two throwaway objects for each of the ~200k records
@@ -742,7 +743,7 @@ class BamRecord {
                 }
             }
             if (len & 1) {
-                seq += SEQRET_DECODER[(ba[p + nPairs] & 0xf0) >> 4];
+                seq += seqAlphabet_ts_1.SEQRET_DECODER[(ba[p + nPairs] & 0xf0) >> 4];
             }
         }
         else {
@@ -754,7 +755,7 @@ class BamRecord {
                 pairs[j] = SEQRET_PAIR_CODES[ba[p + j]];
             }
             if (len & 1) {
-                out[len - 1] = SEQRET_CODES[(ba[p + nPairs] & 0xf0) >> 4];
+                out[len - 1] = seqAlphabet_ts_1.SEQRET_CODES[(ba[p + nPairs] & 0xf0) >> 4];
             }
             seq = textDecoder.decode(out.subarray(0, len));
         }
@@ -809,13 +810,79 @@ class BamRecord {
         if (idx < this.seq_length) {
             const sb = this._byteArray[this.seqStart + (idx >> 1)];
             return (idx & 1) === 0
-                ? SEQRET_DECODER[(sb & 0xf0) >> 4]
-                : SEQRET_DECODER[sb & 0x0f];
+                ? seqAlphabet_ts_1.SEQRET_DECODER[(sb & 0xf0) >> 4]
+                : seqAlphabet_ts_1.SEQRET_DECODER[sb & 0x0f];
         }
         else {
             return undefined;
         }
     }
+    /**
+     * Bind reference bases for this read to resolve its substitutions against,
+     * for a read with no MD tag. `BamFile` calls this for you when it was given a
+     * `fetchReferenceSequence`; call it yourself when you have the bases from
+     * somewhere else.
+     *
+     * **`ref` must cover the whole read**, and this throws if it does not. The
+     * reason is that records are cached and shared between queries (ADR 0006), so
+     * a binding that varied by query would make one query's reads answer out of
+     * another's region. A region covering the read is the same data whichever
+     * query fetched it, so binding it is safe; a partial one is not, and the
+     * per-call `forEachMismatch(cb, {ref})` is where that belongs.
+     */
+    setReference(ref) {
+        if (ref !== undefined && !(0, reference_ts_1.referenceCovers)(ref, this.start, this.end)) {
+            throw new Error(`reference region ${ref.start}-${ref.start + ref.length} does not cover the record at ${this.start}-${this.end}. Records are shared between queries, so only a region covering the whole read can be bound to one; pass a partial region per call instead, as forEachMismatch(cb, {ref})`);
+        }
+        this._reference = ref;
+    }
+    /** the region {@link setReference} bound, if any */
+    get reference() {
+        return this._reference;
+    }
+    /**
+     * Report each difference between this read and the reference —
+     * substitutions, insertions, deletions, reference skips and clips — without
+     * allocating an object per difference. This is the intended way to read a
+     * record's differences; deriving them from `CIGAR` and `MD` yourself takes
+     * rather more of the format to interpret correctly (see {@link Mismatch}).
+     *
+     * Substitutions come from the MD tag when the read has one, and otherwise
+     * from comparing SEQ against reference bases — which have to come from
+     * {@link setReference}, `opts.ref`, or the file's `fetchReferenceSequence`.
+     * With neither MD nor reference, indels and clips are still reported in full
+     * and substitutions are not reported at all: nothing in the record says where
+     * they are.
+     *
+     * @param callback called as
+     *   `(code, refPos, length, bases, qual, refBaseCode, clipLength)`
+     * @param opts optional reference window to restrict to, an `origin` the
+     *   reported positions are relative to, and an optional per-call reference
+     *   region; see {@link MismatchOptions}
+     */
+    forEachMismatch(callback, opts) {
+        (0, mismatches_ts_1.forEachMismatchNumeric)(this.NUMERIC_CIGAR, this.NUMERIC_SEQ, this.seq_length, this.NUMERIC_MD, this.qual, opts?.ref ?? this._reference, this.start, opts?.start ?? Number.NEGATIVE_INFINITY, opts?.end ?? Number.POSITIVE_INFINITY, opts?.origin ?? 0, callback);
+    }
+    /**
+     * The same differences {@link forEachMismatch} reports, as an array of
+     * {@link Mismatch} objects. Convenient; allocates one object per difference,
+     * so the callback form is the one to reach for on a hot path.
+     */
+    getMismatches(opts) {
+        const out = [];
+        this.forEachMismatch((code, refPos, length, bases, qual, refBaseCode, clipLength) => {
+            out.push({
+                code,
+                refPos,
+                length,
+                bases,
+                qual,
+                refBaseCode,
+                clipLength,
+            });
+        }, opts);
+        return out;
+    }
     // Most public BamRecord fields are getters on the prototype, so
     // Object.keys(this) wouldn't include them — JSON.stringify needs an explicit
     // list. Returns the meaningful BAM-spec fields. Return type is widened so