pandora_lib_promethion/src/collection/bam.rs

use std::{
    collections::HashMap,
    fs::{self, File},
    path::PathBuf,
};

use anyhow::{anyhow, Context};
use chrono::{DateTime, Utc};
use dashmap::DashMap;
use glob::glob;
use log::{debug, info, warn};
use rand::{rng, Rng};
use rayon::prelude::*;
use rust_htslib::bam::{ext::BamRecordExtensions, record::Cigar, Read};
use serde::{Deserialize, Serialize};

use crate::config::Config;

use super::flowcells::FlowCell;

#[derive(Debug, Clone, Deserialize, Serialize)]
pub struct WGSBam {
    pub id: String,
    pub time_point: String,
    pub reference_genome: String,
    // pub bam_type: BamType,
    pub path: PathBuf,
    pub modified: DateTime<Utc>,
    pub bam_stats: WGSBamStats,
    // pub cramino: Option<CraminoRes>,
    pub composition: Vec<(String, String, f64)>, // acquisition id, fn
}

// #[derive(Debug, PartialEq, Clone, Deserialize, Serialize)]
// pub enum BamType {
//     WGS,
//     Panel(String),
//     ChIP(String),
// }
//
impl WGSBam {
    pub fn new(path: PathBuf) -> anyhow::Result<Self> {
        let stem = path
            .clone()
            .file_stem()
            .context(format!("Can't parse stem from {}", path.display()))?
            .to_string_lossy()
            .to_string();
        let stem: Vec<&str> = stem.split('_').collect();

        if stem.len() != 3 {
            return Err(anyhow!("Error in bam name: {}", path.display()));
        }

        let id = stem[0].to_string();
        let time_point = stem[1].to_string();

        let reference_genome = stem
            .last()
            .context("Can't get last from stem {stem}")?
            .to_string();

        // let bam_type = if stem.len() == 4 {
        //     match stem[2] {
        //         "oncoT" => BamType::Panel("oncoT".to_string()),
        //         "H3K27ac" => BamType::ChIP("H3K27ac".to_string()),
        //         "H3K4me3" => BamType::ChIP("H3K4me3".to_string()),
        //         _ => return Err(anyhow!("Error in bam name: {}", path.display())),
        //     }
        // } else {
        //     BamType::WGS
        // };
        let file_metadata = fs::metadata(&path)?;
        let modified = file_metadata.modified()?;

        let tp_dir = path
            .parent()
            .context("Can't parse parent from: {bam_path}")?;

        let json_path = format!(
            "{}/{id}_{time_point}_{reference_genome}_info.json",
            tp_dir.to_string_lossy()
        );
        let json_file = PathBuf::from(&json_path);

        if json_file.exists() && json_file.metadata()?.modified()? > modified {
            match Self::load_json(&json_path) {
                Ok(s) => return Ok(s),
                Err(e) => {
                    warn!("Error while loading {}.\n{e}", json_path);
                }
            }
        }

        // let cramino_path = format!(
        //     "{}/{id}_{time_point}_{reference_genome}_cramino.txt",
        //     tp_dir.to_string_lossy()
        // );

        // let cramino = if bam_type == BamType::WGS {
        //     if !PathBuf::from_str(&cramino_path)?.exists() {
        //         // return Err(anyhow!("Cramino file missing {cramino_path}"));
        //         Cramino::default().with_bam(path.to_str().unwrap())?;
        //     }
        //     let mut cramino = Cramino::default().with_result_path(&cramino_path);
        //     cramino
        //         .parse_results()
        //         .context(format!("Error while parsing cramino for {cramino_path}"))?;
        //
        //     if let Some(cramino) = cramino.results {
        //         Some(cramino)
        //     } else {
        //         return Err(anyhow!("Cramino results parsing failed"));
        //     }
        // } else {
        //     None
        // };

        let composition = bam_compo(path.to_string_lossy().as_ref(), 20_000).context(format!(
            "Error while reading BAM composition for {}",
            path.display()
        ))?;

        let s = Self {
            path,
            bam_stats: WGSBamStats::new(&id, &time_point, Config::default())?,
            // cramino,
            id: id.to_string(),
            time_point: time_point.to_string(),
            // bam_type,
            reference_genome,
            composition,
            modified: modified.into(),
        };
        s.save_json(&json_path)?;
        Ok(s)
    }

    pub fn load_json(path: &str) -> anyhow::Result<Self> {
        let f = File::open(path)?;
        let s: Self = serde_json::from_reader(f)?;
        Ok(s)
    }

    pub fn save_json(&self, path: &str) -> anyhow::Result<()> {
        let f = File::create(path)?;
        serde_json::to_writer(f, self)?;
        Ok(())
    }
}

#[derive(Debug, Default, Clone)]
pub struct BamCollection {
    pub bams: Vec<WGSBam>,
}

impl BamCollection {
    pub fn new(result_dir: &str) -> Self {
        load_bam_collection(result_dir)
    }

    pub fn by_acquisition_id(&self) -> HashMap<String, Vec<&WGSBam>> {
        let mut acq: HashMap<String, Vec<&WGSBam>> = HashMap::new();
        for bam in self.bams.iter() {
            for (acq_id, _, _) in bam.composition.iter() {
                if let Some(entry) = acq.get_mut(acq_id) {
                    entry.push(bam);
                } else {
                    acq.insert(acq_id.to_string(), vec![bam]);
                }
            }
        }
        acq
    }

    pub fn get(&self, id: &str, time_point: &str) -> Vec<&WGSBam> {
        self.bams
            .iter()
            .filter(|b| b.id == id && b.time_point == time_point)
            .collect()
    }

    pub fn by_id_completed(&self, min_diag_cov: f32, min_mrd_cov: f32) -> Vec<WGSBam> {
        self.bams
            .iter()
            // .filter(|b| matches!(b.bam_type, BamType::WGS))
            .filter(|b| match b.time_point.as_str() {
                "diag" => b.bam_stats.global_coverage >= min_diag_cov as f64,
                "mrd" => b.bam_stats.global_coverage >= min_mrd_cov as f64,
                _ => false,
            })
            // .filter(|b| match &b.cramino {
            //     Some(cramino) => match b.time_point.as_str() {
            //         "diag" => cramino.mean_length >= min_diag_cov as f64,
            //         "mrd" => cramino.mean_length >= min_mrd_cov as f64,
            //         _ => false,
            //     },
            //     _ => false,
            // })
            .cloned()
            .collect()
    }

    pub fn ids(&self) -> Vec<String> {
        let mut ids: Vec<String> = self.bams.iter().map(|b| b.id.clone()).collect();
        ids.sort();
        ids.dedup();
        ids
    }
}

pub fn load_bam_collection(result_dir: &str) -> BamCollection {
    let pattern = format!("{}/*/*/*.bam", result_dir);
    let bams = glob(&pattern)
        .expect("Failed to read glob pattern")
        // .par_bridge()
        .filter_map(|entry| {
            match entry {
                Ok(path) => match WGSBam::new(path) {
                    Ok(bam) => return Some(bam),
                    Err(e) => warn!("{e}"),
                },
                Err(e) => warn!("Error: {:?}", e),
            }
            None
        })
        .collect();

    BamCollection { bams }
}

pub fn bam_compo(file_path: &str, sample_size: usize) -> anyhow::Result<Vec<(String, String, f64)>> {
    let mut bam = rust_htslib::bam::Reader::from_path(file_path)?;

    let mut rgs: HashMap<( String, String ), u64> = HashMap::new();
    for result in bam.records().filter_map(Result::ok).take(sample_size) {
        if let ( rust_htslib::bam::record::Aux::String(s),  rust_htslib::bam::record::Aux::String(b)) = ( result.aux(b"RG")?, result.aux(b"fn")? ) {
            *rgs.entry(( s.to_string(), b.to_string())).or_default() += 1;
        }
    }

    Ok(rgs
        .into_iter()
        .map(|(k, n)| (k.0.to_string(), k.1.to_string(), n as f64 * 100.0 / sample_size as f64))
        .map(|(rg, f, p)| {
            let (a, _) = rg.split_once('_').unwrap();
            let (b, _) = f.split_once('_').unwrap();
            (a.to_string(), b.to_string(), p)
        })
        .collect())
}

pub fn bam_has_fc(bam: &WGSBam, flow_cell: &FlowCell) -> anyhow::Result<bool> {
    let mut bam = rust_htslib::bam::Reader::from_path(&bam.path)?;

    let mut rgs: HashMap<String, u64> = HashMap::new();
    for result in bam.records().filter_map(Result::ok).take(2_000) {
        if let rust_htslib::bam::record::Aux::String(s) = result.aux(b"fn")? {
            *rgs.entry(s.to_string()).or_default() += 1;
        }
    }

    for k in rgs.keys() {
        if k.contains(&flow_cell.sample_sheet.flow_cell_id) {
            return Ok(true);
        }
    }

    Ok(false)
}

pub struct BamReadsSampler {
    pub positions: Vec<(String, u64)>,
    pub reader: rust_htslib::bam::IndexedReader,
    current_index: usize,
}

impl BamReadsSampler {
    pub fn new(path: &str, n: usize) -> anyhow::Result<Self> {
        debug!("Reading BAM file: {path}");
        let reader = rust_htslib::bam::IndexedReader::from_path(path)?;
        let header = reader.header();

        let contig_len: Vec<(String, u64)> = header
            .target_names()
            .into_iter()
            .map(|target_name| {
                let tid = header.tid(target_name).unwrap();
                (
                    String::from_utf8(target_name.to_vec()).unwrap(),
                    header.target_len(tid).unwrap(),
                )
            })
            .collect();

        let positions = sample_random_positions(&contig_len, n);

        Ok(Self {
            positions,
            reader,
            current_index: 0,
        })
    }
}

impl Iterator for BamReadsSampler {
    type Item = Result<rust_htslib::bam::Record, rust_htslib::errors::Error>;

    fn next(&mut self) -> Option<Self::Item> {
        loop {
            if self.current_index < self.positions.len() {
                let (contig, pos) = &self.positions[self.current_index];

                match self.reader.fetch((contig, *pos, pos + 1)) {
                    Ok(_) => (),
                    Err(e) => warn!("Error while reading BAM {e}"),
                }

                let result = self.reader.records().next();

                self.current_index += 1;
                if result.is_some() {
                    return result;
                }
            } else {
                return None;
            }
        }
    }
}

pub fn n_mapped_reads(bam_path: &str) -> anyhow::Result<u64> {
    let mut bam = rust_htslib::bam::IndexedReader::from_path(bam_path)?;
    let mapped = bam
        .index_stats()?
        .into_iter()
        .filter(|(tid, _, _, _)| *tid < 24)
        .map(|(_, _, m, _)| m)
        .sum();
    Ok(mapped)
}

// 0-based inclusif
pub fn nt_pileup(
    bam: &mut rust_htslib::bam::IndexedReader,
    chr: &str,
    position: u32,
    with_next_ins: bool,
) -> anyhow::Result<Vec<u8>> {
    use rust_htslib::{bam, bam::Read};
    let mut bases = Vec::new();
    bam.fetch((chr, position, position + 1))?;
    let mut bam_pileup = Vec::new();
    for p in bam.pileup() {
        let pileup = p.context(format!(
            "Can't pileup bam at position {}:{} (0-based)",
            chr, position
        ))?;
        let cur_position = pileup.pos();
        if cur_position == position {
            for alignment in pileup.alignments() {
                match alignment.indel() {
                    bam::pileup::Indel::Ins(_len) => bam_pileup.push(b'I'),
                    bam::pileup::Indel::Del(_len) => bam_pileup.push(b'D'),
                    _ => {
                        let record = alignment.record();
                        if record.seq_len() > 0 {
                            if let Some(b) = base_at(&record, position, with_next_ins)? {
                                bases.push(b);
                            }
                        } else if alignment.is_del() {
                            bases.push(b'D');
                        }
                    }
                }
            }
        }
    }
    Ok(bases)
}

pub fn base_at(
    record: &rust_htslib::bam::record::Record,
    at_pos: u32,
    with_next_ins: bool,
) -> anyhow::Result<Option<u8>> {
    let cigar = record.cigar();
    let seq = record.seq();
    let pos = cigar.pos() as u32;

    let mut read_i = 0u32;
    // let at_pos = at_pos - 1;
    let mut ref_pos = pos;
    if ref_pos > at_pos {
        return Ok(None);
    }

    for (id, op) in cigar.iter().enumerate() {
        let (add_read, add_ref) = match *op {
            Cigar::Match(len) | Cigar::Equal(len) | Cigar::Diff(len) => (len, len),
            Cigar::Ins(len) => (len, 0),
            Cigar::Del(len) => (0, len),
            Cigar::RefSkip(len) => (0, len),
            Cigar::SoftClip(len) => (len, 0),
            Cigar::HardClip(_) | Cigar::Pad(_) => (0, 0),
        };
        // If at the end of the op len and next op is Ins return I
        if with_next_ins && ref_pos + add_read == at_pos + 1 {
            if let Some(Cigar::Ins(_)) = cigar.get(id + 1) {
                return Ok(Some(b'I'));
            }
        }

        if ref_pos + add_ref > at_pos {
            // Handle deletions directly
            if let Cigar::Del(_) = *op {
                return Ok(Some(b'D'));
            } else if let Cigar::RefSkip(_) = op {
                return Ok(None);
            } else {
                let diff = at_pos - ref_pos;
                let p = read_i + diff;
                return Ok(Some(seq[p as usize]));
            }
        }

        read_i += add_read;
        ref_pos += add_ref;
    }
    Ok(None)
}

pub fn counts_at(
    bam: &mut rust_htslib::bam::IndexedReader,
    chr: &str,
    position: u32,
) -> anyhow::Result<HashMap<String, i32>> {
    let p = nt_pileup(bam, chr, position, false)?
        .iter()
        .map(|e| String::from_utf8(vec![*e]).unwrap())
        .collect::<Vec<_>>();
    let mut counts = HashMap::new();
    for item in p.iter() {
        *counts.entry(item.to_string()).or_insert(0) += 1;
    }
    Ok(counts)
}

pub fn ins_pileup(
    bam: &mut rust_htslib::bam::IndexedReader,
    chr: &str,
    position: u32,
) -> anyhow::Result<Vec<String>> {
    let mut bases = Vec::new();
    bam.fetch((chr, position, position + 10))?;
    for p in bam.pileup() {
        let pileup = p.context(format!(
            "Can't pileup bam at position {}:{} (0-based)",
            chr, position
        ))?;
        let cur_position = pileup.pos();
        // Ins in the next position
        if cur_position == position + 1 {
            // debug!("{cur_position}");
            for alignment in pileup.alignments() {
                let record = alignment.record();
                if record.seq_len() > 0 {
                    if let Some(b) = ins_at(&record, position)? {
                        bases.push(b);
                    }
                }
            }
        }
    }
    Ok(bases)
}

pub fn ins_at(
    record: &rust_htslib::bam::record::Record,
    at_pos: u32,
) -> anyhow::Result<Option<String>> {
    use rust_htslib::bam::record::Cigar;

    let cigar = record.cigar();
    let seq = record.seq();
    let pos = cigar.pos() as u32;

    let mut read_i = 0u32;
    let mut ref_pos = pos;
    if ref_pos > at_pos {
        return Ok(None);
    }
    // debug!(
    //     "read: {}",
    //     String::from_utf8(record.qname().to_vec()).unwrap()
    // );

    for op in cigar.iter() {
        let (add_read, add_ref) = match *op {
            Cigar::Match(len) | Cigar::Equal(len) | Cigar::Diff(len) => (len, len),
            Cigar::Ins(len) => (len, 0),
            Cigar::Del(len) => (0, len),
            Cigar::RefSkip(len) => (0, len),
            Cigar::SoftClip(len) => (len, 0),
            Cigar::HardClip(_) | Cigar::Pad(_) => (0, 0),
        };

        if ref_pos + add_read > at_pos && ref_pos + add_read < at_pos + 10 {
            if let Cigar::Ins(v) = *op {
                // debug!(
                //     "ins size {v} @ {} (corrected {})",
                //     ref_pos + add_read,
                //     (ref_pos + add_read) - v - 1
                // );

                if (ref_pos + add_read) - v - 1 == at_pos {
                    let inserted_seq =
                        seq.as_bytes()[read_i as usize..(read_i + v) as usize].to_vec();
                    return Ok(Some(String::from_utf8(inserted_seq)?));
                }
            }
        }

        read_i += add_read;
        ref_pos += add_ref;
    }
    Ok(None)
}

pub fn counts_ins_at(
    bam: &mut rust_htslib::bam::IndexedReader,
    chr: &str,
    position: u32,
) -> anyhow::Result<HashMap<String, i32>> {
    let p = ins_pileup(bam, chr, position)?;
    let mut counts = HashMap::new();
    for item in p.iter() {
        *counts.entry(item.to_string()).or_insert(0) += 1;
    }
    Ok(counts)
}

pub fn sample_random_positions(chromosomes: &Vec<(String, u64)>, n: usize) -> Vec<(String, u64)> {
    let mut rng = rng();
    let total_length: u64 = chromosomes.iter().map(|(_, length)| length).sum();

    (0..n)
        .map(|_| {
            let random_position = rng.random_range(0..total_length);
            let mut cumulative_length = 0;

            for (chr, length) in chromosomes {
                cumulative_length += length;
                if random_position < cumulative_length {
                    let position_within_chr = random_position - (cumulative_length - length);
                    return (chr.clone(), position_within_chr);
                }
            }

            unreachable!("Should always find a chromosome")
        })
        .collect()
}

#[derive(Debug, Clone, Deserialize, Serialize)]
pub struct WGSBamStats {
    pub all_records: u128,
    pub n_reads: u128,
    pub mapped_yield: u128,
    pub global_coverage: f64,
    pub karyotype: Vec<(i32, u64, String, u128)>,
    pub n50: u128,
    pub by_lengths: Vec<(u128, u32)>,
}

impl WGSBamStats {
    pub fn new(id: &str, time: &str, config: Config) -> anyhow::Result<Self> {
        let bam_path = config.solo_bam(id, time);
        let Config {
            bam_min_mapq,
            bam_n_threads,
            ..
        } = config;
        let mut bam = rust_htslib::bam::Reader::from_path(bam_path)?;
        let h = bam.header().clone();
        let header = rust_htslib::bam::Header::from_template(&h);
        bam.set_threads(bam_n_threads as usize)?;

        let mut mapped_lengths = Vec::new();
        let mut all_records = 0;
        let mut n_reads = 0;
        let mut lengths_by_tid: HashMap<i32, Vec<u128>> = HashMap::new();

        for read in bam
            .rc_records()
            .map(|x| x.expect("Failure parsing Bam file"))
            .inspect(|_| all_records += 1)
            .filter(|read| {
                read.flags()
                    & (rust_htslib::htslib::BAM_FUNMAP
                        | rust_htslib::htslib::BAM_FSECONDARY
                        | rust_htslib::htslib::BAM_FQCFAIL
                        | rust_htslib::htslib::BAM_FDUP) as u16
                    == 0
            })
            .filter(|r| r.mapq() >= bam_min_mapq)
        {
            n_reads += 1;
            let mapped_len = (read.reference_end() - read.pos()) as u128;
            mapped_lengths.push(mapped_len);
            lengths_by_tid
                .entry(read.tid())
                .or_default()
                .push(mapped_len);
            if n_reads % 500_000 == 0 {
                info!("{n_reads} reads parsed");
            }
        }

        let mapped_yield = mapped_lengths.par_iter().sum::<u128>();
        let genome = get_genome_sizes(&header)?;
        let genome_size = genome.values().map(|v| *v as u128).sum::<u128>();
        let global_coverage = mapped_yield as f64 / genome_size as f64;

        let by_lengths: DashMap<u128, u32> = DashMap::new();
        mapped_lengths
            .par_iter()
            .for_each(|l| *by_lengths.entry(*l).or_default() += 1);

        let mut by_lengths: Vec<(u128, u32)> =
            by_lengths.iter().map(|e| (*e.key(), *e.value())).collect();
        by_lengths.par_sort_by(|a, b| a.0.cmp(&b.0));
        // This is not n50
        let n50 = by_lengths[by_lengths.len() / 2].0;

        let mut karyotype: Vec<(i32, u64, String, u128)> = lengths_by_tid
            .iter()
            .map(|(tid, lengths)| {
                let contig = String::from_utf8(h.tid2name(*tid as u32).to_vec()).unwrap();
                (
                    *tid,
                    *genome.get(&contig).unwrap(),
                    contig,
                    lengths.par_iter().sum::<u128>(),
                )
            })
            .collect();

        karyotype.sort_by(|(ac, _, _, _), (bc, _, _, _)| ac.cmp(bc));

        Ok(Self {
            all_records,
            n_reads,
            mapped_yield,
            global_coverage,
            karyotype,
            n50,
            by_lengths,
        })
    }

    pub fn print(&self) {
        println!("N records\t{}", self.all_records);
        println!("N counted reads\t{}", self.n_reads);

        println!("Mapped yield [Gb]\t{:.2}", self.mapped_yield as f64 / 1e9);
        println!("Mean coverage\t{:.2}", self.global_coverage);
        self.karyotype
            .iter()
            .for_each(|(_, contig_len, contig, contig_yield)| {
                println!(
                    "{contig}\t{:.2}",
                    (*contig_yield as f64 / *contig_len as f64) / self.global_coverage
                );
            });
    }
}

pub fn get_genome_sizes(header: &rust_htslib::bam::Header) -> anyhow::Result<HashMap<String, u64>> {
    let mut genome = HashMap::new();
    for (key, records) in header.to_hashmap() {
        for record in records {
            if key == "SQ" {
                genome.insert(
                    record["SN"].to_string(),
                    record["LN"]
                        .parse::<u64>()
                        .expect("Failed parsing length of chromosomes"),
                );
            }
        }
    }
    Ok(genome)
}

/// Result of querying a read at a reference position.
#[derive(Clone, Debug, Eq, PartialEq)]
pub enum PileBase {
    A,
    C,
    G,
    T,
    N,
    Ins,                 // insertion immediately after the queried base
    Del((Vec<u8>, u32)), // deletion covering the queried base
    Skip,                // reference skip (N)
}

/// Decode one HTSlib 4bit nucleotide (0=A,1=C,2=G,3=T,4=N, …) to `Base`
#[inline]
fn decode(b: u8) -> PileBase {
    match b & 0b1111 {
        0 => PileBase::A,
        1 => PileBase::C,
        2 => PileBase::G,
        3 => PileBase::T,
        _ => PileBase::N,
    }
}

pub fn base_at_new(
    record: &rust_htslib::bam::Record,
    at_pos: i64,
    with_next_ins: bool,
) -> anyhow::Result<Option<PileBase>> {
    if record.pos() > at_pos {
        return Ok(None);
    }

    let mut ref_pos = record.pos();
    let mut read_pos = 0_i64;
    let cigar = record.cigar();
    let seq = record.seq().as_bytes(); // already expanded to ASCII (A,C,G,T,N)

    for (i, op) in cigar.iter().enumerate() {
        let (consume_r, consume_ref) = match op {
            Cigar::Match(l) | Cigar::Equal(l) | Cigar::Diff(l) => (*l as i64, *l as i64),
            Cigar::Ins(l) | Cigar::SoftClip(l) => (*l as i64, 0),
            Cigar::Del(l) | Cigar::RefSkip(l) => (0, *l as i64),
            _ => (0, 0),
        };

        /* look ahead for an insertion immediately AFTER the queried base */
        if with_next_ins
            && ref_pos + consume_ref == at_pos + 1
            && matches!(cigar.get(i + 1), Some(Cigar::Ins(_)))
        {
            return Ok(Some(PileBase::Ins));
        }

        /* Does the queried reference coordinate fall inside this CIGAR op? */
        if ref_pos + consume_ref > at_pos {
            return Ok(Some(match op {
                Cigar::Del(e) => PileBase::Del((record.qname().to_vec(), *e)),
                Cigar::RefSkip(_) => PileBase::Skip,
                _ => {
                    let offset = (at_pos - ref_pos) as usize;
                    if read_pos as usize + offset >= seq.len() {
                        return Ok(None); // out‑of‑range, malformed BAM
                    }
                    decode(seq[read_pos as usize + offset])
                }
            }));
        }

        read_pos += consume_r;
        ref_pos += consume_ref;
    }

    Ok(None) // beyond alignment
}

/// Return the pile‑up of **all reads** at `chr:pos` (0‑based, inclusive).
///
/// * `bam`           – an open [`rust_htslib::bam::IndexedReader`].  
/// * `chr` / `pos`   – reference contig name and coordinate.  
/// * `with_next_ins` – if `true`, report [`Base::Ins`] when an insertion starts
///                     **right after** the queried base.
///
/// The function bounds the internal pile‑up depth to 10 000 reads to protect
/// against malformed BAM files that could otherwise allocate unbounded memory.
///
/// # Errors
///
/// Propagates any I/O or parsing error from *rust‑htslib*, wrapped in
/// [`anyhow::Error`].
pub fn nt_pileup_new(
    bam: &mut rust_htslib::bam::IndexedReader,
    chr: &str,
    pos: u32,
    with_next_ins: bool,
) -> anyhow::Result<Vec<PileBase>> {
    // Storage for the per‑read observations.
    let mut pile = Vec::new();

    // Restrict BAM iterator to the one‑base span of interest.
    bam.fetch((chr, pos, pos + 1))?;

    // Hard cap on depth (adjust if your use‑case needs deeper coverage).
    bam.pileup().set_max_depth(10_000);

    // Stream the pileup; HTSlib walks the reads for us.
    for pileup in bam.pileup() {
        // Attach context to any low‑level error.
        let pileup = pileup.context(format!("can't pile up BAM at {chr}:{pos}"))?;

        // Skip other positions quickly.
        if pileup.pos() != pos {
            continue;
        }

        for aln in pileup.alignments() {
            // Handle the three indel states reported by HTSlib.
            match aln.indel() {
                rust_htslib::bam::pileup::Indel::Ins(_) => {
                    pile.push(PileBase::Ins); // insertion *starts at* this base
                }
                rust_htslib::bam::pileup::Indel::Del(e) => {
                    let qname = aln.record().qname().to_vec();
                    pile.push(PileBase::Del((qname, e))); // deletion length > 1, but still covers here
                }
                rust_htslib::bam::pileup::Indel::None => {
                    // Regular match/mismatch: delegate to `base_at`.
                    if let Some(base) = base_at_new(&aln.record(), pos as i64, with_next_ins)? {
                        pile.push(base);
                    }
                }
            }
        }
    }
    Ok(pile)
}